|Mehmet Kesimer, PhD, Research Assistant Professor of Biochemistry and Biophysics|
One of the main focuses of my work is the characterization of the large mucin gene products (Mr 2-3 million) and the complexes they make (Mr 10-100 million) essential for the formation of the mucus gels vital for epithelial protection and function. My current work is focused around the human lung where there are many hypersecretory human diseases including asthma, cystic fibrosis, and chronic bronchitis in which these glycoconjugates are centrally implicated. My proteomic interest includes designing and implementing novel mass spectral methods for identifying proteins and glycoproteins in mucus gels where conventional proteomic methods do not work. Many of the molecules are heavily modified and thus MS methods for characterizing post-translational modifications are vital in this work. We are using state-of-the-art mass spectrometer systems such as MALDI-TOF and 1D, 2D and even 3D separations employing nano-UPLC-MS-MS.
Mucins and Mucin Interactome in the Airway
Normal airway mucus is highly structured, containing five dominant mucins (MUC5AC, MUC5B, MUC1, MUC4 and MUC16) an excess of 100 proteins that form distinct protein complexes many of which are centered around these mucins. These complexes constitute a discrete secretory entity we call the ‘mucin interactome’. We propose that gel properties emerge from a dynamic interplay between the mucins and proteins and that these structures engender specific rheological properties of mucus that tune it to removal by cough and flow clearance.
|Figure 1. MUC5B (gel/mucus forming mucin).|
The Effect of Cystic Fibrosis Post-Secretory Settings on Mucin Expansion and Maturation
Cystic Fibrosis in the lung is characterized by the failure of mucus clearance. Clearly mucus plays an important role in all manifestations of CF. There are some two hundred’ mainly globular proteins in human sputum, but it is the gel-forming mucins MUC5AC and MUC5B that we believe create the structural scaffold. The mechanisms underpinning the packaging of the large gel-forming mucins in granules and their subsequent expansion to mature the mucus are very poorly understood. This work is investigating the hypothesis that in the absence of CFTR, mucin expansion/mucus maturation is impaired due to different post-secretory environment in CF mucus. In testing this hypothesis, using the resolving power of the electron microscope in addition to a broad range of biochemical and biophysical techniques, we are testing: 1) To determine the factors effecting the normal maturation of the MUC5B just after secretion; 2) To test the hypothesis that the MUC5B mucin as born into the CF environment is prejudiced in its development and subsequent maturation.
Funding: CF foundation (KESIME10I0)
Role of Human Airway Epithelium Derived Exosome-Like Vesicles in Innate Defense of the Lung
Exosomes are small, 50-150 nm, organelles secreted by different cell types including epithelial, haematopoietic and some tumour cells. The molecular organisation of these structures depends mainly on the cellular source from which they are derived. The detailed roles of the exosomes remain largely unexplained. We have demonstrated their presence in lung secretions and propose that they play important roles in innate lung defence, such as binding to and neutralizing the viruses. The objective of this work is to elucidate the organization of the human tracheobronchial epithelial (HTBE) exosomes and identify their cellular origin, ultimately we wish to understand their roles in airway host defense. Our preliminary data from HTBE cell culture indicate that exosomes are highly organized structures arising from different cell types to be found in the culture and that they interact with human influenza virus. Many of them have membrane tethered mucin coats that control their recognition properties.
Funding: American Lung Association (RG-167538-N)
Ongoing Research Support
KESIME10I0 Kesimer (PI) 04/01/2010-03/31-2012 Cystic Fibrosis Foundation
Effect of CF-post-secretory settings on mucin expansion and maturation
The major goals of this project is to understand why mucus clearance is problematic in the CF lung. The concept underlying this proposal is that the environment in the CF airway impairs "mucin expansion/maturation" ("unpacking to form mucus gels") after secretion from granular forms of MUC5B.
RG-167538-N Kesimer (PI) 07/01/2010-06/30/2012 American Lung Association
"The role of human airway epithelium derived exosomes in innate defense of the lung"
The major goals of this project is to further elucidate the organization of the HTB exosomes, identify their cellular origin and assess their role in airway host defense against respiratory viral and/or bacterial pathogens and to determine the nature of HTBE exosomes-influenza virus interactions.
1 R01 HL103940-01 Kesimer (PI) 07/01/2010 - 06/30/2015 NIH, NHLBI (Pending administrative review)
“The role of mucin-protein interactions in the innate defense of the lung”
The major scientific goal of this proposal is to elucidate the organization of the airway gel layer, which is formed by very large glycoproteins termed mucins (MUC5B and MUC5AC) coupled with other protective biomolecules through labile interactions, and assess the role of these interactions in airway mucus integrity as an essential pre-requisite for therapeutic intervention.
5 P50 HL 084934-02 Boucher (PI) 9/15/06-7/31/11 NIH/NHLBI
SCCOR in Host Factors in Chronic Lung Disease, Project I: The Mucus Transport Apparatus: Why Two Layers?
The major goals of this project are, using a human bronchial epithelial cell line and normal and pathological sputum, to characterize the properties of the peri-ciliary gel layer and to identify the molecular mechanisms underlying the MUC5AC-rich gel and the MUC5B flowing mucus.
For a more exhaustive list of publications, please see links from the Pubmed feed in the righthand column.
Mehmet Kesimer, Alexander M. Makhov, Jack D. Griffith, Pedro Verdugo and John K. Sheehan. Unpacking a gel forming mucin: a view of MUC5B organization after granular release. Am J Physiol Lung Cell Mol Physiol 298: L15–L22, 2010
Kesimer M, Kilic N, Mehrotra R, Thornton DJ, and Sheehan JK. Identification of salivary mucin MUC7 binding proteins from Streptococcus gordonii. BMC Microbiol 9: 163, 2009.
Mehmet Kesimer, Margaret Scull, Brian Brighton, Genevieve DeMaria, Kimberlie Burns, Wanda O’Neal, Raymond J. Pickles and John K. Sheehan. “Characterization of exosome-like vesicles released from human tracheobronchial ciliated epithelium: a possible role in innate defense. FASEB J 23: 1858-1868, 2009.
Mehmet Kesimer, Sara Kirkham, Raymond Pickles, Ashley Henderson, Neil Alexis, Genevieve DeMaria, David Knight, David Thornton, and John Sheehan. "Tracheobronchial air-liquid interface cell culture: a model for innate mucosal defense of the upper airways?" Am J Physiol Lung Cell Mol Physiol. 2009 Jan;296(1):L92-L100.
Mehmet Kesimer and John K. Sheehan. Analyzing the functions of large glycoconjugates through the dissipative properties of their absorbed layers using the gel-forming mucin MUC5B as an example. Glycobiology. 2008 Jun;18(6):463-72.
John K. Sheehan, Mehmet Kesimer and Raymond Pickles. Chapter: Innate immunity and mucus structure and function, in: Innate Immunity to Pulmonary Infection, Novartis Foundation series No 279, ISBN: 0-470-02656-1, pages 155-69. (2006).
The University of North Carolina at Chapel Hill
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